1. Field of the Invention
"This 371 application claims the benefit of PCT/FR94/00422, filed Nov. 10, 1994, which claims the benefit of French application FR93/05125, filed Apr. 30, 1993."
The present invention relates to new recombinant viruses, to their preparation and to their use in gene therapy, for the transfer and expression in vivo of desired genes. More specifically, it relates to new recombinant viruses comprising an inserted gene whose expression in vivo makes it possible to regulate the plasma levels of apolipoproteins. The present invention also relates to pharmaceutical compositions comprising the said recombinant viruses. More particularly, the present invention relates to defective recombinant viruses and to their use for the prevention or treatment of pathologies linked to dyslipoproteinemias which are known for their serious consequences at the cardiovascular and neurological level.
2. Description of Related Art
Dyslipoproteinemias are disorders of the metabolism of the lipoproteins responsible for the transport, in the blood and peripheral fluids, of lipids such as cholesterol and triglycerides. They result in major pathologies, linked respectively to hypercholesterolemia or hypertriglyceridemia, such as especially atherosclerosis. Atherosclerosis is a polygenic complex disease which is defined from the histological point of view by deposits (lipid or fibrolipid plaques) of lipids and of other blood derivatives in the wall of the large arteries (aorta, coronary arteries, carotid). These plaques, which are calcified to a greater or lesser extent according to the progression of their process, can be associated with lesions and are linked to the accumulation, in the arteries, of fatty deposits consisting essentially of cholesterol esters. These plaques are accompanied by a thickening of the arterial wall, with hypertrophy of the smooth muscle, the appearance of spumous cells and the accumulation of fibrous tissue. The atheromatous plaque is very clearly in relief on the wall, which confers on it a stenosing character responsible for vascular occlusions by atheroma, thrombosis or embolism which occur in the patients most affected. Hypercholosterolemias can therefore result in very serious cardiovascular pathologies such as infarct, sudden death, cardiac decompensation, cerebral vascular accidents and the like.
It is therefore particularly important to be able to have available treatments which make it possible to reduce, in certain pathological situations, the plasma cholesterol levels or even to stimulate the efflux of cholesterol (reverse transport of the cholesterol) in the peripheral tissues in order to discharge the cells having accumulated cholesterol within the context of the formation of an atheroma plaque. The cholesterol is carried in the blood by various lipoproteins including the low-density lipoproteins (LDL) and the high-density lipoproteins (HDL). The LDLs are synthesized in the liver and make it possible to supply the peripheral tissues with cholesterol. In contrast, the HDLs capture cholesterol in the peripheral tissues and transport it to the liver where it is stored and/or degraded.
At present, dyslipemias and in particular hypercholesterolemias are treated essentially by means of compounds which act either on the biosynthesis of cholesterol (inhibitors of hydroxymethylglutaryl-coenzymeA reductase, statins), or on the capture and elimination of bile cholesterol (sequestrants or resins), or alternatively on lipolysis by a mode of action which remains to be elucidated from the molecular point of view (fibrates). Consequently, all the major categories of drugs which have been used in this indication (sequestrants, fibrates or statins), are designed only for the preventive aspect of the formation of the atheroma plaque and not in fact for the treatment of the atheroma. The current treatment for atheroma, following a coronary accident, are only palliative since they do not act on cholesterol homeostasis and they are surgical acts (coronary by-pass, angioplasty).
The present invention constitutes a new therapeutic approach for the treatment of pathologies linked to dyslipoproteinemias. It proposes an advantageous solution to the disadvantages of the prior art, by demonstrating the possibility of treating pathologies linked to dyslipoproteinemias by gene therapy, by the transfer and expression in vivo of genes capable of regulating the plasma levels of apolipoproteins. The invention thus offers a simple means permitting a specific and effective treatment of these pathologies.
Gene therapy consists in correcting a deficiency or an abnormality (mutation, aberrant expression and the like) by introduction of a genetic information into the cell or affected organ. This genetic information can be introduced either in vitro into a cell extracted from the organ, the modified cell then being reintroduced into the body, or directly in vivo into the appropriate tissue. In this second case, various techniques exist, of which various techniques of transfection involving complexes of DNA and DEAE-dextran (Pagano et al., J. Virol. 1 (1967) 891), of DNA and nuclear proteins (Kaneda et al., Science 243 (1989) 375), of DNA and lipids (Felgner et al., PNAS 84 (1987) 7413), the use of liposomes (Fraley et al., J. Biol. Chem. 255 (1980) 10431), and the like. More recently, the use of viruses as vectors for the transfer of genes appeared as a promising alternative to these physical techniques of transfection. In this respect, various viruses were tested for their capacity to infect certain cellular populations, in particular the retroviruses (RSV, KMS, MKS, and the like), the HSV viruses, the adeno-associated viruses and the adenoviruses.